Department of Chemistry, University of Southern California, Los Angeles, CA 90089.
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115.
Proc Natl Acad Sci U S A. 2022 Feb 8;119(6). doi: 10.1073/pnas.2120287119.
A variety of signals, including inflammasome activation, trigger the formation of large transmembrane pores by gasdermin D (GSDMD). There are primarily two functions of the GSDMD pore, to drive lytic cell death, known as pyroptosis, and to permit the release of leaderless interleukin-1 (IL-1) family cytokines, a process that does not require pyroptosis. We are interested in the mechanism by which the GSDMD pore channels IL-1 release from living cells. Recent studies revealed that electrostatic interaction, in addition to cargo size, plays a critical role in GSDMD-dependent protein release. Here, we determined computationally that to enable electrostatic filtering against pro-IL-1β, acidic lipids in the membrane need to effectively neutralize positive charges in the membrane-facing patches of the GSDMD pore. In addition, we predicted that salt has an attenuating effect on electrostatic filtering and then validated this prediction using a liposome leakage assay. A calibrated electrostatic screening factor is necessary to account for the experimental observations, suggesting that ion distribution within the pore may be different from the bulk solution. Our findings corroborate the electrostatic influence of IL-1 transport exerted by the GSDMD pore and reveal extrinsic factors, including lipid and salt, that affect the electrostatic environment.
多种信号，包括炎性小体激活，触发gasdermin D（GSDMD）形成大的跨膜孔。GSDMD 孔主要有两个功能，一是驱动裂解细胞死亡，称为细胞焦亡，二是允许无领导者白细胞介素-1（IL-1）家族细胞因子的释放，这个过程不需要细胞焦亡。我们感兴趣的是 GSDMD 孔通道将 IL-1 从活细胞中释放的机制。最近的研究表明，除了货物大小外，静电相互作用在 GSDMD 依赖性蛋白释放中起着关键作用。在这里，我们通过计算确定，为了使静电过滤能够对抗 pro-IL-1β，膜中的酸性脂质需要有效地中和 GSDMD 孔膜面向片段中的正电荷。此外，我们预测盐对静电过滤有衰减作用，然后使用脂质体渗漏测定法验证了这一预测。需要校准的静电筛选因子来解释实验观察结果，这表明离子在孔内的分布可能与体相溶液不同。我们的发现证实了 GSDMD 孔对 IL-1 转运的静电影响，并揭示了影响静电环境的外在因素，包括脂质和盐。
